Browsing by Author "Palta, Priit"
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Item A molecular tool for menstrual cycle phase dating of endometrial samples in endometriosis transcriptome studies(2019) Saare, Merli; Merli, Triin; Teder, Hindrek; Paluoja, Priit; Palta, Priit; Koel, Mariann; Kirss, Fred; Karro, Helle; Sõritsa, Deniss; Salumets, Andres; Krjutškov, Kaarel; Peters, MaireTranscriptome profiling of 57 endometrial receptivity genes specifies the menstrual cycle phase of endometrial samples.Item Computational methods for DNA copy number detection(2015-09-16) Palta, PriitDNA koopiaarvu variantideks või muutusteks nimetatakse selliseid erinevusi inimeste geneetilises materjalis, mille puhul mingi DNA lõigu koopiaarv on erinev oodatavast koopiaarvust kaks (üks koopia mingit kindlat DNA järjestust emalt päritud kromosoomil ja üks koopia isalt päritud kromosoomil). DNA koopiate vähenemist nimetatakse deletsiooniks ning vastavaid DNA variante nimetatakse deletsioonideks. DNA koopiate juurdetulemist nimetatakse duplitseerumiseks ning selliseid kahest suurema koopiaarvuga variante vastavalt duplikatsioonideks. Antud doktoritöös uuriti inimese DNA koopiaarvu variante, nende seotust erinevate haigustega ja nende tekkimise ja pärandumise eripärasid. Kasutades DNA mikrokiipe ehk geenikiipe uuriti esmalt kas ja millised DNA koopiaarvu muutused võivad olla seotud vaimse arengu mahajäämusega (VAM-ga). Uurides perekondasid, kus ühel või mitmel liikmel oli diagnoositud VAM, leiti mitmeid juba varem VAM-ga seostatud DNA koopiaarvu muutusi ning lisaks leiti ka mitmeid uusi DNA koopiaarvu variante, mille esinemine võib olla seotud VAM-e väljakujunemisega. Sarnane uuring viidi läbi ka korduva spontaanse raseduse katkemise probleemiga paaride ja naiste puhul. Võrreldes nende patsientide gruppi kuuluvate naiste DNA koopiaarvu muutusi ning nende sagedusi terveid emasid sisaldavate kontroll-grupi indiviidide omadega, leiti statistiliselt ja bioloogiliselt oluline erinevus muutunud koopiaarvuga DNA lõigus, mis sisaldab PDZD2 ja GOLPH3 geene ja kus esinevate duplikatsioonide „omamine“ suurendas naistel märkimisväärselt spontaanse raseduse katkemise ohtu. Doktoritöö viimases osas uuriti Tartu Ülikooli Eesti Geenivaramu ja rahvusvahelise HapMap projekti poolt kogutud tõsiste haigusteta inimestel esinevaid DNA koopiaarvu muutusi ja nende pärandumist perekondades. Selle uuringu üheks huvitavamaks tulemuseks oli deletsioonide alapärandumine vanematelt lastele ehk deletsioone kandvaid DNA regioone esines laste genoomides oluliselt vähem, kui normaalse Mendeliaalse (juhusliku) pärandumise korral oleks oodata võinud. Uurides duplikatsioonide regioone perekondades leiti aga, et kaks kolmandikku duplikatsioonides esinevatest DNA koopiatest ei olnud identsed (üksteise täpsed koopiad), vaid mõnevõrra erinevad, demonstreerides seniajani teadmata olnud alleelse varieeruvuse määra DNA duplikatsioonide regioonides.Item Creating basis for introducing non‐invasive prenatal testing in the Estonian public health setting(John Wiley & Sons, Ltd., 2019-11) Žilina, Olga; Rekker, Kadri; Kaplinski, Lauris; Sauk, Martin; Paluoja, Priit; Teder, Hindrek; Ustav, Eva‐Liina; Tõnisson, Neeme; Reimand, Tiia; Ridnõi, Konstantin; Palta, Priit; Vermeesch, Joris Robert; Krjutškov, Kaarel; Kurg, Ants; Salumet, AndresObjective The study aimed to validate a whole‐genome sequencing‐based NIPT laboratory method and our recently developed NIPTmer aneuploidy detection software with the potential to integrate the pipeline into prenatal clinical care in Estonia. Method In total, 424 maternal blood samples were included. Analysis pipeline involved cell‐free DNA extraction, library preparation and massively parallel sequencing on Illumina platform. Aneuploidies were determined with NIPTmer software, which is based on counting pre‐defined per‐chromosome sets of unique k‐mers from sequencing raw data. SeqFF was implemented to estimate cell‐free fetal DNA (cffDNA) fraction. Results NIPTmer identified correctly all samples of non‐mosaic trisomy 21 (T21, 15/15), T18 (9/9), T13 (4/4) and monosomy X (4/4) cases, with the 100% sensitivity. However, one mosaic T18 remained undetected. Six false‐positive (FP) results were observed (FP rate of 1.5%, 6/398), including three for T18 (specificity 99.3%) and three for T13 (specificity 99.3%). The level of cffDNA of <4% was estimated in eight samples, including one sample with T13 and T18. Despite low cffDNA level, these two samples were determined as aneuploid. Conclusion We believe that the developed NIPT method can successfully be used as a universal primary screening test in combination with ultrasound scan for the first trimester fetal examination.Item TAC-seq: targeted DNA and RNA sequencing for precise biomarker molecule counting(2018) Teder, Hindrek; Koel, Mariann; Paluoja, Priit; Jatsenko, Tatjana; Rekker, Kadri; Laisk-Podar, Triin; Kukuškina, Viktorija; Velthut-Meikas, Agne; Fjodorova, Olga; Peters, Maire; Kere, Juha; Salumets, Andres; Palta, Priit; Krjutškov, KaarelTargeted next-generation sequencing (NGS) methods have become essential in medical research and diagnostics. In addition to NGS sensitivity and high-throughput capacity, precise biomolecule counting based on unique molecular identifier (UMI) has potential to increase biomolecule detection accuracy. Although UMIs are widely used in basic research its introduction to clinical assays is still in progress. Here, we present a robust and cost-effective TAC-seq (Targeted Allele Counting by sequencing) method that uses UMIs to estimate the original molecule counts of mRNAs, microRNAs, and cell-free DNA. We applied TAC-seq in three different clinical applications and compared the results with standard NGS. RNA samples extracted from human endometrial biopsies were analyzed using previously described 57 mRNA-based receptivity biomarkers and 49 selected microRNAs at different expression levels. Cell-free DNA aneuploidy testing was based on cell line (47,XX, +21) genomic DNA. TAC-seq mRNA profiling showed identical clustering results to transcriptome RNA sequencing, and microRNA detection demonstrated significant reduction in amplification bias, allowing to determine minor expression changes between different samples that remained undetermined by standard NGS. The mimicking experiment for cell-free DNA fetal aneuploidy analysis showed that TAC-seq can be applied to count highly fragmented DNA, detecting significant (p = 7.6 × 10−4) excess of chromosome 21 molecules at 10% fetal fraction level. Based on three proof-of-principle applications we demonstrate that TAC-seq is an accurate and highly potential biomarker profiling method for advanced medical research and diagnostics.